Final year Bpharm 7th semester
Vikrant Ashok Shirke
Gsmcop, Wagholi, Pune
BP701T - INSTRUMENTAL METHODS OF ANALYSIS.
UNIT-II (16-44 Marks)
Syllabus:-
FTIR Spectroscopy:- Introduction, fundamental modes of vibrations in poly atomic
molecules, samples handling, factors affecting vibrations.
Instrumentation:- Sources of radiation, wavelength selectors.
Detectors:- Golay cell, Bolometer, thermocouple, thermister, pyroelectric detectors.
FTIR instrument, sample handling attechments:- DRS and ATR and applications.
Flame Photometry:- Principle, interferences, instrumentation and applications.
Atomic absorption spectroscopy:- Principle, interference, instrumentation and
applications.
Nepheloturbidimetry:- Introduction.
Q.1. Attempt the following:- (3 Marks Each)
1) Explain the types of molecular vibrations in IR spectroscopy?
Classify Vibrational modes in IR spectroscopy?
★ Classification of Molecular vibrations/modes in IR spectroscopy:-
In Infrared (IR) Spectroscopy, molecular vibrations are central to how molecules absorb
infrared light. These vibrations occur when atoms in a molecule move relative to one
another while the molecule remains intact.
➢ Types of Molecular Vibrations.
1) Stretching Vibrations These involve changes in the bond length between two atoms.
➔ Types of Streching:-
a) Symmetric Stretching:- Atoms move simultaneously in and out along the bond axis.
[ Example: In a CO₂ molecule, both C=O bonds stretch or compress together. ]
b) Asymmetric Stretching:- One bond shortens while the other lengthens.
Occurs at a higher frequency than symmetric stretching.
2) Bending Vibrations These involve changes in the bond angle between atoms.
➔ Types of bending:-
a) Scissoring:- Two atoms move toward and away from each other like a pair of scissors.
b) Rocking:- Atoms move in the same direction, like a pendulum swinging side to side.
c) Wagging:- Two atoms move up and down out of the plane, in phase.
d) Twisting:- One atom moves up while the other moves down out of the plane.
2) Discuss the applications of flames Photometry?
★ Applications of Flame Photometry
, 1) Flame Photometry, also known as Flame Atomic Emission Spectroscopy, is a simple,
rapid, and cost-effective technique primarily used to determine the concentration of
certain metal ions.
2) Used to measure concentrations of sodium (Na⁺), potassium (K⁺), calcium (Ca²⁺), and
lithium (Li⁺) in blood serum, urine, and plasma.
3) Helps in diagnosing electrolyte imbalances,kidney function, cardiac health.
4) Quantitative determination of alkali and alkaline earth metals in drug formulations.
5) Ensures compliance with pharmacopeial standards.
6) Determination of essential nutrients like potassium, sodium, and calcium in soil and
fertilizers.
7) Estimation of mineral content in food products, such as Milk, fruit juices, vegetables, and
salt.
8) Detection of metal contaminants (like Na⁺, K⁺, Ca²⁺) in natural waters, industrial effluents,
and wastewater.
9) Monitors pollution levels and assesses water quality.
10)Determination of metallic impurities in chemical reagents and raw materials.
3) Write the principle involved in nepheloturbidometry?
★ Principle of Nepheloturbidometry:
1) Nepheloturbidometry is based on the scattering of light by suspended particles in a
solution. When a beam of light passes through a solution containing fine, insoluble
particles (such as a precipitate), the light is scattered in various directions due to the
interaction with the particles.
2) Nephelometry measures the intensity of scattered light, typically at an angle (usually 90°)
to the incident beam.
3) Turbidometry measures the reduction in intensity of the transmitted light due to scattering
and absorption by the particles.
4) The amount of light scattered (or the decrease in transmitted light) is proportional to the
concentration of the suspended particles in the sample.
5) The intensity of scattered or transmitted light is directly related to the number and size of
the particles present in the solution.
6) This technique is commonly used for:
a) Measuring protein levels (e.g, in immunoassays).
b) Determining sulfate, chloride, or other precipitating ions.
c) Water and wastewater analysis for turbidity levels.
4) Explain the sample handling in IR spectroscopy?
Explain DRS and ATR Technique of Sample Handling in IR Spectroscopy? (5 marks)
★ Sample Handling in IR Spectroscopy
1) Solid Samples:
a) KBr Pellet Method: The solid sample is finely ground with dry potassium bromide (KBr),
which is IR-transparent.
The mixture is compressed into a thin, transparent pellet using a pellet press.
This is the most common method for recording IR spectra of solids.
b) Mull Technique: The solid is ground and mixed with a mulling agent (like liquid paraffin)
to form a thick paste (mull).
This paste is sandwiched between two salt plates (e.g, NaCl) and placed in the IR beam.
, c) Diffuse Reflectance: The powdered sample is mixed with an IR-transparent matrix (like
KBr) and analyzed using a special reflectance accessory, Useful for minimal sample
preparation and surface analysis.
2) Liquid Samples:
a) Liquid Cell Method: A drop of the liquid is placed between two IR-transparent salt plates
(NaCl or KBr) to form a thin film.
The cell is mounted in the IR spectrophotometer.
For volatile liquids, sealed cells may be used to prevent evaporation.
b) Solvent Selection: Solvents used must not absorb in the IR region or should have minimal
interference (e.g, carbon tetrachloride or chloroform for organic samples).
3) Gaseous Samples:
a) Gas Cell: Gases are analyzed using long-path gas cells (typically 5–10 cm or longer), fitted
with IR-transparent windows, The sample is introduced into the cell, and spectra are
recorded.
1) Diffuse Reflectance Spectroscopy (DRS)
➢ Principle: DRS is based on the diffuse reflection of IR radiation from a powdered or rough
solid sample.
When IR light strikes a rough surface, part of it is absorbed, and the rest is scattered in
various directions.
This scattered light carries information about the sample’s vibrational transitions.
➢ Working: 1) The powdered sample is mixed with a non-absorbing material like KBr or KCl.
2) IR radiation is directed at the sample.
3) The diffusely reflected light is collected using ellipsoidal mirrors and sent to the detector.
➢ Instrumentation:
Uses a special diffuse reflectance accessory.
The sample is placed in a cup or holder designed for powders.
➢ Applications:
1) Analysis of powders, opaque solids, or inorganic materials.
2) Non-destructive testing, Useful in pharmaceutical, mineral, and polymer analysis.
➢ Advantages:
1) Minimal sample preparation.
2) Suitable for insoluble and non-transparent materials.
4) Attenuated Total Reflectance (ATR)
➢ Principle: ATR works on the principle of total internal reflection.
An IR beam enters a crystal with high refractive index (e.g, ZnSe, diamond) and reflects
internally at the interface where the sample is in contact.
An evanescent wave penetrates a few microns into the sample and gets absorbed depending on
the sample's vibrational modes.
➢ Working: 1) The sample is placed in direct contact with the ATR crystal.
2) IR light enters the crystal and undergoes multiple internal reflections.
3) At each reflection, a small part of the wave interacts with the sample, absorbing IR energy at
specific frequencies.
4) The remaining light is collected and analyzed.
➢ Instrumentation:
Includes an ATR accessory fitted with an IR-transparent crystal.
Sample is pressed against the crystal using a clamp.
Vikrant Ashok Shirke
Gsmcop, Wagholi, Pune
BP701T - INSTRUMENTAL METHODS OF ANALYSIS.
UNIT-II (16-44 Marks)
Syllabus:-
FTIR Spectroscopy:- Introduction, fundamental modes of vibrations in poly atomic
molecules, samples handling, factors affecting vibrations.
Instrumentation:- Sources of radiation, wavelength selectors.
Detectors:- Golay cell, Bolometer, thermocouple, thermister, pyroelectric detectors.
FTIR instrument, sample handling attechments:- DRS and ATR and applications.
Flame Photometry:- Principle, interferences, instrumentation and applications.
Atomic absorption spectroscopy:- Principle, interference, instrumentation and
applications.
Nepheloturbidimetry:- Introduction.
Q.1. Attempt the following:- (3 Marks Each)
1) Explain the types of molecular vibrations in IR spectroscopy?
Classify Vibrational modes in IR spectroscopy?
★ Classification of Molecular vibrations/modes in IR spectroscopy:-
In Infrared (IR) Spectroscopy, molecular vibrations are central to how molecules absorb
infrared light. These vibrations occur when atoms in a molecule move relative to one
another while the molecule remains intact.
➢ Types of Molecular Vibrations.
1) Stretching Vibrations These involve changes in the bond length between two atoms.
➔ Types of Streching:-
a) Symmetric Stretching:- Atoms move simultaneously in and out along the bond axis.
[ Example: In a CO₂ molecule, both C=O bonds stretch or compress together. ]
b) Asymmetric Stretching:- One bond shortens while the other lengthens.
Occurs at a higher frequency than symmetric stretching.
2) Bending Vibrations These involve changes in the bond angle between atoms.
➔ Types of bending:-
a) Scissoring:- Two atoms move toward and away from each other like a pair of scissors.
b) Rocking:- Atoms move in the same direction, like a pendulum swinging side to side.
c) Wagging:- Two atoms move up and down out of the plane, in phase.
d) Twisting:- One atom moves up while the other moves down out of the plane.
2) Discuss the applications of flames Photometry?
★ Applications of Flame Photometry
, 1) Flame Photometry, also known as Flame Atomic Emission Spectroscopy, is a simple,
rapid, and cost-effective technique primarily used to determine the concentration of
certain metal ions.
2) Used to measure concentrations of sodium (Na⁺), potassium (K⁺), calcium (Ca²⁺), and
lithium (Li⁺) in blood serum, urine, and plasma.
3) Helps in diagnosing electrolyte imbalances,kidney function, cardiac health.
4) Quantitative determination of alkali and alkaline earth metals in drug formulations.
5) Ensures compliance with pharmacopeial standards.
6) Determination of essential nutrients like potassium, sodium, and calcium in soil and
fertilizers.
7) Estimation of mineral content in food products, such as Milk, fruit juices, vegetables, and
salt.
8) Detection of metal contaminants (like Na⁺, K⁺, Ca²⁺) in natural waters, industrial effluents,
and wastewater.
9) Monitors pollution levels and assesses water quality.
10)Determination of metallic impurities in chemical reagents and raw materials.
3) Write the principle involved in nepheloturbidometry?
★ Principle of Nepheloturbidometry:
1) Nepheloturbidometry is based on the scattering of light by suspended particles in a
solution. When a beam of light passes through a solution containing fine, insoluble
particles (such as a precipitate), the light is scattered in various directions due to the
interaction with the particles.
2) Nephelometry measures the intensity of scattered light, typically at an angle (usually 90°)
to the incident beam.
3) Turbidometry measures the reduction in intensity of the transmitted light due to scattering
and absorption by the particles.
4) The amount of light scattered (or the decrease in transmitted light) is proportional to the
concentration of the suspended particles in the sample.
5) The intensity of scattered or transmitted light is directly related to the number and size of
the particles present in the solution.
6) This technique is commonly used for:
a) Measuring protein levels (e.g, in immunoassays).
b) Determining sulfate, chloride, or other precipitating ions.
c) Water and wastewater analysis for turbidity levels.
4) Explain the sample handling in IR spectroscopy?
Explain DRS and ATR Technique of Sample Handling in IR Spectroscopy? (5 marks)
★ Sample Handling in IR Spectroscopy
1) Solid Samples:
a) KBr Pellet Method: The solid sample is finely ground with dry potassium bromide (KBr),
which is IR-transparent.
The mixture is compressed into a thin, transparent pellet using a pellet press.
This is the most common method for recording IR spectra of solids.
b) Mull Technique: The solid is ground and mixed with a mulling agent (like liquid paraffin)
to form a thick paste (mull).
This paste is sandwiched between two salt plates (e.g, NaCl) and placed in the IR beam.
, c) Diffuse Reflectance: The powdered sample is mixed with an IR-transparent matrix (like
KBr) and analyzed using a special reflectance accessory, Useful for minimal sample
preparation and surface analysis.
2) Liquid Samples:
a) Liquid Cell Method: A drop of the liquid is placed between two IR-transparent salt plates
(NaCl or KBr) to form a thin film.
The cell is mounted in the IR spectrophotometer.
For volatile liquids, sealed cells may be used to prevent evaporation.
b) Solvent Selection: Solvents used must not absorb in the IR region or should have minimal
interference (e.g, carbon tetrachloride or chloroform for organic samples).
3) Gaseous Samples:
a) Gas Cell: Gases are analyzed using long-path gas cells (typically 5–10 cm or longer), fitted
with IR-transparent windows, The sample is introduced into the cell, and spectra are
recorded.
1) Diffuse Reflectance Spectroscopy (DRS)
➢ Principle: DRS is based on the diffuse reflection of IR radiation from a powdered or rough
solid sample.
When IR light strikes a rough surface, part of it is absorbed, and the rest is scattered in
various directions.
This scattered light carries information about the sample’s vibrational transitions.
➢ Working: 1) The powdered sample is mixed with a non-absorbing material like KBr or KCl.
2) IR radiation is directed at the sample.
3) The diffusely reflected light is collected using ellipsoidal mirrors and sent to the detector.
➢ Instrumentation:
Uses a special diffuse reflectance accessory.
The sample is placed in a cup or holder designed for powders.
➢ Applications:
1) Analysis of powders, opaque solids, or inorganic materials.
2) Non-destructive testing, Useful in pharmaceutical, mineral, and polymer analysis.
➢ Advantages:
1) Minimal sample preparation.
2) Suitable for insoluble and non-transparent materials.
4) Attenuated Total Reflectance (ATR)
➢ Principle: ATR works on the principle of total internal reflection.
An IR beam enters a crystal with high refractive index (e.g, ZnSe, diamond) and reflects
internally at the interface where the sample is in contact.
An evanescent wave penetrates a few microns into the sample and gets absorbed depending on
the sample's vibrational modes.
➢ Working: 1) The sample is placed in direct contact with the ATR crystal.
2) IR light enters the crystal and undergoes multiple internal reflections.
3) At each reflection, a small part of the wave interacts with the sample, absorbing IR energy at
specific frequencies.
4) The remaining light is collected and analyzed.
➢ Instrumentation:
Includes an ATR accessory fitted with an IR-transparent crystal.
Sample is pressed against the crystal using a clamp.
